There has hitherto been a microparticle detection device that counts microparticles developed in liquid or on a membrane or a slide glass or conducts a property inspection on the microparticles by irradiating the microparticles with light and detecting fluorescence or scattered light generated from the microparticles. Here, the microparticles include inorganic particles, microorganisms, cells, red blood cells, white blood cells, and platelets in the blood, endothelial cells, small cell debris of the above tissues, and microparticles in the blood. When the microparticles are present in the liquid, they serve as a microparticle suspension.
A flow cytometer is popular as a detection method for the microparticles. In this flow cytometer, a suspension of the microparticles is run through a capillary together with sheath fluid. A part of the capillary is irradiated with laser light, and the kind of the microparticles and the size of the microparticles are sorted by detecting scattered light or fluorescence generated when the microparticles are irradiated with the light. For example, by labeling particles with a fluorescent reagent that bonds to specific particles, the number of fluorescence-emitting particles can be counted and only the specific particles can be counted. This flow cytometer can detect particles from the submicron order to about 10 μm, and can achieve high-accuracy detection.
However, the above-described flow cytometer that can measure even particles of the submicron order is a large-sized and expensive system.
In contrast, there is a particle detection method that measures an image of particles and identifies the kind and size of the particles from information about the image. Since this method performs detection and analysis of the particles by using the image, it is sometimes called an image cytometer in contrast to the flow cytometer. Imaging methods include photographing using a microscope and a digital camera and a method that performs imaging by detecting scattered light or fluorescence while two-dimensionally scanning an optical head.
In the case of photographing using the microscope and the digital camera, when particles have a size of 1 μm or more, a high-accuracy image can be measured. However, when particles of the submicron order are measured, a microscope having a high-power objective lens and a highly sensitive (that is, low-noise and wide dynamic range) digital camera are necessary. Hence, the system is considerably expensive. In the case of submicron particles, since the light wavelength and the particle size are equal, imaging performance is reduced by the diffraction limit, and it is difficult to exactly identify the particle size.
Further, particles can be easily detected by using a fluorescence microscope system as the above microscope. However, similarly, the light wavelength and the particle size are equal to each other when the particles are submicron particles. Hence, the particle size cannot be identified exactly. Further, since fluorescence from the microparticles is faint, a highly sensitive digital camera is necessary.
In contrast to this, in the system that detects scattered light or fluorescence while scanning the optical head, laser light from the optical head is collected and applied to particles, and the optical head is two-dimensionally scanned to perform imaging while detecting scattered light or fluorescence generated from the particles.
As such a system that detects light while scanning the optical head, Japanese Patent No. 3928846 (PTL 1) discloses a scanner having a confocal optical system.
In the scanner having the confocal optical system disclosed in PTL 1, laser light emitted from a laser excitation light source is turned into parallel light by a collimator lens, passes through a hole of a holed mirror and a first lens in an optical head, and enters a sample set on a sample stage that is movable in the X-direction and the Y-direction. When the sample is a fluorescence sample, a fluorescent substance is excited by the laser light and fluorescence is emitted. When the sample is a storage phosphor sheet, a stimulable phosphor is excited by the laser light, and stimulable light is emitted. The light thus emitted from the sample is turned into parallel light by the first lens, is reflected by a portion around the hole of the holed mirror, and is collected by a second lens. Then, the light passes through a confocal switch member disposed at the focal position of the second lens, and is photoelectrically detected by a photomultiplier, so that analog data is generated.
In the system that thus detects light while scanning the optical head relative to the sample, when submicron particles are detected, the irradiation spot diameter of the laser light is equal to or larger than the particle size. For this reason, individual particles are not resolved in an image obtained as a result of two-dimensional scanning. Hence, it is difficult to directly measure the size of the particles from the image. However, even when the irradiation spot size of the laser light is larger than the particle size, the intensity of scattered light generated from the particles differs according to the particle size. Hence, it is possible to identify the particle size from the intensity of scattered light. This is because the particle size and the intensity of scattered light are correlated with each other.
In this case, although a detector for detecting scattered light with high sensitivity (a low-noise and wide dynamic range detector) and a laser light source are necessary, it is possible to configure a system that is less expensive than the system using the microscope having the high-power objective lens and the highly sensitive digital camera.
However, the above-described conventional system, which detects light while scanning the optical head relative to the sample, has the following problems.
That is, in the above system that detects scattered light from the sample by two-dimensional scanning, measurement needs to be performed while decreasing the scan pitch of the two-dimensional scanning as the particle size decreases. This is because particles may be skipped when the scan pitch is larger than the particle diameter. However, a new problem occurs, that is, the measurement time increases as the measurement pitch (scan pitch) decreases.
When the irradiation spot diameter of laser light is set to be larger than the particle size, the particles are not skipped. However, the intensity of scattered light decreases and the particle size may be misread.
However, the scanner having the confocal optical system disclosed in PTL 1 is a typical fluorescent imaging system, but is not intended to detect microparticles. Therefore, PTL 1 does not describe the problem caused when the scan pitch is decreased because the particle size is small and the problem caused when the irradiation spot diameter of laser light is increased.